Fluorescent Proteins: FRET
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Collection
...expression level and maturation time, and other environmental factors. When designing your experiment, be ...
We narrowed to 682 results for: RON
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CRISPR References and Information
TypeCollection...construct ; pMA122 (negative selection marker); pGH8 (neuronal co-injection marker); pCFJ104 (body wall muscle... -
CRISPR History and Development for Genome Engineering
TypeCollection...through non-homologous end joining (NHEJ) , an error-prone process that produces insertions/deletions disrupting... -
Viral Vectors 101: The AAV Retrograde Serotype
TypeBlog Post...these regions, a process driven by neurons. By studying how neurons are physically connected, we can begin...these connections in the brain are (drum roll) neurons! Neurons are really cool because they can transmit ...There are two methods of neuronal tracing (Fig. 1): one to visualize neurons that send information to ... functionally dissect specific neurons within their natural environment, all while reciting their favorite...defined population of neurons. For example, say I wanted to access all the neurons that signaled from the...ligands activates both neurons at the injection site and any projection neurons retrogradely transduced... efficient retrograde access to projection neurons." Neuron 92.2 (2016): 372-382. PubMed PMID: 27720486... -
FLEx Technology and Optogenetics: Flipping the switch on gene expression with high spatial and temporal resolution
TypeBlog Post.... What do microbial rhodopsins and neurons have in common? Neuronal stimulations are triggered by the ...the neuron fires, sending the electrical signal down the axon (Lodish et al., 2000). Most neurons in the...Vandenberghe, 2019). How to achieve strong opsin expression in specific neuronal cell types then? This is where... 2014). How does FLEx enable strong opsin expression in specific neuronal cells? This can be achieved ...enabling its expression under a strong promoter only in specific neuronal cells. FLEx ensures, therefore...et al. "Silencing neurons: tools, applications, and experimental constraints." Neuron 95.3 (2017): 504-...What is optogenetics? Optogenetics: Controlling neurons with light In 2005, a team from Stanford University... -
New Tools Enable CRISPRa for Neuroscience Applications
TypeBlog Post...specify and assemble diverse arrays of neurons, forming the neuronal circuits that will later be refined ...individual neuronal populations. The Yang Lab’s AAV plasmids used to express Cre under neuron-specific ...kinase II alpha (aCaMKII; excitatory neurons) and Synapsin-1 (pan-neuronal) promoters (Figure 1b), and sgRNAs...response to neuronal activity (hence, “immediate-early”) and consequently regulates neuronal physiology...between Bdnf I-overexpressing neurons and Bdnf IV-overexpressing neurons. Among the upregulated genes ...contributions of transcript isoforms to neuronal physiology. The Day Lab’s neuron-optimized lentiviral vectors can...process. However, while the neuronal marker TUJ1 is useful for visualizing neuronal differentiation by immunohistochemistry... -
Viral Vectors 101: Systemic Capsids
TypeBlog Post...CNS: neurons and astrocytes Mice No (LY6C1) Hanlon et al., 2019 AAV-BI-28 CNS: neurons, astrocytes...astrocytes, but its neuron transduction encompasses more molecularly-defined neuronal subtypes and with ...and is highly specific for neurons, with less bias toward inhibitory neurons than AAV-PHP.eB (Fig. 4c, ...transduce 80% of dorsal root ganglia (DRG) neurons, as well as neurons located in the enteric nervous system...primate nervous systems. Neuron, 110(14), 2242-2257.e6. https://doi.org/10.1016/j.neuron.2022.05.003 Chen, X...-specific labeling. Neuron,109(9), 1449–1464.e13. https://doi.org/10.1016/j.neuron.2021.03.011 Hanlon,...cell type specificity, or retrograde transport in neurons. (b) Genetic cargo can be customized to control... -
Deciphering the Mysteries of Behavior with Viral Vectors
TypeBlog Post...individual neurons and in terms of their interactions with one another. In addition to the neuronal networks... (hM3Dq) into a subset of neurons within the LC of mice. When those neurons are chemogenetically activated...reward circuit, and the medial prefrontal cortex (mPFC) are involved. Neurons from the mPFC project to the...findings by controlling neurons in the mPFC with light. Activation of these neurons via the channel Channelrhodopsin...protein that activates neurons upon light stimulation. When they activated the Drd1 neurons 24 hours before ... "Optogenetic stimulation of medial prefrontal cortex Drd1 neurons produces rapid and long-lasting antidepressant...They also don’t give any clues about the types of neurons involved or whether their involvement is activating... -
AAVs CREATed for Gene Delivery to the CNS and PNS
TypeBlog Post...for genetically manipulating neurons throughout the whole brain. Neurons of the peripheral nervous system...transduced a higher number of neurons overall (GFP+ cells that also stained for a neuronal marker). Check out tables...for? 1. Multi-color labeling of neurons Multi-color labeling of neurons is for more than making pretty ...single-cell anatomical study and neuronal tracing, aka determining where a neuron’s axons and dendrites extend...in order to decrease the number of neurons that get labeled, neurons are more likely to be transduced by...interest, which, in the case of Chan et al, was neurons and astrocytes. Why screen for IV deliverable ...and GFP intensity and transduction efficiency of neurons were measured. On average, AAV.PHP.eB and AAV.PHP.S... -
Announcing the Winners of the 2021 Michael Davidson and Roger Tsien Commemorative Conference Awards
TypeBlog Post...periaqueductal gray (PAG), with red neurons expressing tdTomato and green neurons expressing cFos. Image from ...the activity of parvalbumin (PV)-expressing neurons, neurons that the lab previously found had a role in...light activates the neurons (via oChIEF) and a chemogenetic agonist inhibits the neuron (via hM4D). She fused...on the PV neurons is in progress, she has validated the dual virus method to study CR neurons which are...fluorescence based on the release of glutamate from neurons and this new version of the biosensor is faster... to the collaborative and highly interactive environment that Janelia offers,” says Aggarwal.In addition... in electroporated in rat hippocampal cultured neurons, imaged 14 days post-transfection, excited at 505... -
Channelrhodopsins with improved light sensitivity for minimally-invasive optogenetics
TypeBlog Post...method that lets you fire neurons with the flick of a light switch. Neurons are not typically persuaded...remains inactive and neurons don't fire. With blue light, ions flow into the neuron and causes it to fire...ChRger. In both experiments, neurons expressing ChRgers produced stronger photocurrents at low light intensity...activate a neuron. ChRgers, however, have a higher conductance, so they can still depolarize neurons at lower...dopaminergic neurons in the VTA, this should propel the mice to increase their nose-poking behavior. Strong nose-poking...shines on excitatory channelrhodopsin-expressing neurons, the channelrhodopsins respond by opening and allowing... allowing an influx of ions into the neuron which generates an action potential (Figure 1). This light-induced... -
Viral Vectors 101: Pseudotyping
TypeBlog Post...Genetically Targeted Neurons. Neuron 53:639–647 . https://doi.org/10.1016/j.neuron.2007.01.033 Additional...the previous neuron (Wichersham et al., 2007). However, once rabies dG enters this neuron, it cannot infect...However, pseudotyping allows these viruses to infect neurons. Besides altering the host tropism of your virus...reasons VSV-G is used to transduce stem cells and neurons. However, VSV-G’s broad tropism can be undesired...Rabies glycoprotein The rabies glycoprotein targets neuronal cells and can travel in a retrograde direction...the cell body, making it a great tool to study neuronal connectivity. However, because of rabies virus...Lentiviral vectors are also great at infecting neurons and cause minimal inflammatory response, but are... -
Zebrafish as a Model for Behavior: Swimming into the Optogenetic Spotlight
TypeBlog Post...they make the inside of the neuron more positive or more negative. Neuronal stimulation: Depolarization...depolarization. They therefore function as a neuron “on switch.” Neuronal inhibition: Hyperpolarization Conversely...cord neurons using the Gal4/UAS system. This created fish lines with different groups of neurons that ...activity, enables researchers to silence or incite neuronal firing and study subsequent effects on behavior...cross the cell membrane when exposed to light. Neuronal firing depends on these charged particles and ...Depolarization When the inside of the neuron becomes positive over a certain threshold, it becomes depolarized and...Conversely, when the inside of the neuron becomes more negative it becomes less likely to fire and is said... -
Fluorescent Proteins 101: Photoactivatable Fluorescent Proteins
TypeBlog Post...reversible PA-FP is the naturally occurring Dronpa protein (named after dron, a ninja term for vanishing, and pa...both activates and inactivates Dronpa is not ideal, since this means Dronpa is quickly inactivated and less... to find a version of Dronpa that doesn't become inactivated as quickly. Padron is a monomeric, reversible... mutating Dronpa (T59M / V60A / N94I / P141L / G155S / V157G / M159Y / F190S) (7). Padron’s “positive”...switching behavior is the opposite of Dronpa; when exposed to blue light, Padron is photoconverted from a dark...Emission (nm) Dronpa Monomer UV-Violet Dark/Green N/A (high) High 503/ 518 Padron Monomer Blue Dark...Correlative light and electron microscopy (CLEM) is an imaging technique that combines electron microscopy (EM... -
Hot Plasmids Spring 2024
TypeBlog Post...www.janelia.org/open-science/voltron-and-positron. Both Ace2N-mNeon-A122D and Voltron2 are available as Cre-dependent...33714753. Voltron2 voltage sensors By Brian O’Neill New viral vector preps expressing the Voltron2 voltage...performance. Figure 4: Structure of Voltron or Voltron2 conjugated to a JaneliaFluor dye emitting...HaloTag domain. Find Voltron2 plasmids here! Abdelfattah, A.S., et al. (2023) Neuron, 111(10), 1547–1563... SARS-CoV-2 research New viral vector preps of Voltron2 voltage sensors AAV vectors for targeting microglia... to facilitate SARS-CoV-2 research By Ashley Waldron Ready-to-use recombinant antibodies were recently..., are now available! Building on their earlier Voltron indicators (a fusion of Ace2N and HaloTag, Figure... -
Advanced Uses of Cre-lox and Flp-FRT - A Neuroscientist’s View
TypeBlog Post...Visualizing single neurons in the living brain Figure 1: Plasmid mix to label neuronal morphology (...labeling of cortical neurons and their synapses. How can you label a single neuron and its synapses...DNA’, is derived from the yeast S. cerevisiae (Gronostajski and Sadowski 1985). The FLP system works in ...neuroscience Manipulation of specific cell types in neuronal networks Double inverted vectors expressing light...manipulate specific cell types (populations) within neuronal networks in living animals. To accomplish such...electroporation. To achieve sparse labeling of individual neurons one could reduce the plasmids amounts to reduce...efficiency. However, if the cellular structure of a neuron and its synapse should be visualized at the same... -
15 Hot Plasmids from 2017
TypeBlog Post...linked to disease, including motor neurons (-3mnx1/Hb9), pan neuronal (elavl3/HuC), microglial/macrophage...specific neuronal subtypes was not available until now. In order to target specific neuronal subtypes ...type SOX2 with either a SOX2-VP16 (strong activator) or SOX2-HP1 (strong repressor) fusion protein, the authors...interest. In order to improve the study of specific neuronal cell types involved in neurodegenerative diseases...), and astrocytic (gfap).This toolbox adds new neuronal tools to the expanding gateway compatible vectors...to express the transcription factors from a polycistronic cassette. However, it was unknown whether variations...variations in gene expression from these polycistronic cassettes could lead to comparable experimental... -
Important Considerations in Optogenetics Behavioral Experiments
TypeBlog Post... Inhibition of neuronal activation is a little more complicated since basal neuronal activity in your...advent of optogenetics, ICSS can now be done with neuronal cell-type and temporal precision. In ICSS, a test...actually stimulating or inhibiting the targeted neuronal population prior to behavioral testing. One method...a reliable and widely used molecular marker of neuronal activation and is a sufficient readout to proceed...readout may be necessary to prove inhibition of neuronal activation, but it may still be possible to use...example, as shown in this study [3], hippocampal neurons are activated when the mouse is placed in a novel... the authors were able to show that HR blocked neuronal activation in the novel context as determined ... -
Hot Plasmids May 2018 - Optogenetics, Decaffeination, Biosensors, and Fluorescent Protein Tools
TypeBlog Post...expression in E. coli. This operon is under the transcriptional control of a strong constitutive promoter (...powerful tool used by neuroscientists to interrogate neuronal networks. A major drawback of this method is the... the stray excitation of neighboring neurons in opsin dense regions. To overcome this issue, Shemesh et... C-terminus of the protein. Unlike the diffuse neuronal expression observed with wild type CoChR, soCoChR...describe a novel system for the precise mapping of neuronal networks at the single-cell level. Various soCoChR...PMID: 29184208. Synthetic decaffeination operon Article contributed by Angela Kaczmarczyk Listen...Listen to the synthetic decaffeination operon podcast segment With the help of engineered microbes, used... -
Lighting Up Cell Signaling with Photoswitchable Kinases
TypeBlog Post...cyan light, Dronpa dimers dissociate, Dronpa goes dark, and MEK1 is active or “on.” Dronpa’s color thus...Additionally, dimeric Dronpa is less prone to aggregation in cells compared to the tetrameric Dronpa it was derived... by attaching two dimerizing photoswitchable Dronpa (dron= a ninja term for vanishing, and pa= photoactivation... the Dronpa dimers interact and lock MEK1 in an “off” setting by blocking the active site. Dronpa also...similarity. For these three kinases, photoswitchable Dronpa domains were inserted at sites homologous to the...here. In addition, it could be possible to use pdDronpa to control other kinases and proteins. Zhou et...proteins could potentially be created by inserting pdDronpa domains into loops of a protein of interest. If...